Over two thirds of spinal cord injury (SCI) patients have anatomical preservation at the injury site, yet this preserved tissue is typically completely or partially dysfunctional. Therefore, to improve function in these patients, strategies are needed that enhance the function of the remaining connections by enabling the inherent plasticity of the CNS. We found an unanticipated interplay between a neurotrophin and cellular immune processes that may provide a means to promote plasticity after SCI. Moreover, we have evidence that this interplay may be exploited to induce neuroplasticity in chronic SCI. Our earlier work showed that unilateral viral-vector mediated over-expression of Neurotrophin-3 (NT-3) in lumbar motoneurons induced axon growth from the contralateral corticospinal tract (CST) towards the source of the NT-3. Subsequently, we showed that there is an immune component to the NT-3-induced axonal sprouting and that microglia are likely involved. This proposal will determine the role that microglia play in supporting post-SCI axonal growth and determine the molecular mechanisms involved. We hypothesize that NT-3 re-programs microglia that become activated during Wallerian degeneration, which then induce axonal sprouting of surviving CST axons. We have 3 aims that are the sum of close collaborations between SCI research laboratories of Baylor College of Medicine and the Ohio State University. In the first aim we will define the microglial-derived factors elicited by NT-3 that can stimulate axonal growth. Preliminary data show that NT-3 re-programs inflammatory macrophages and microglia to a less inflammatory state. We propose to use unbiased RNA-seq to identify candidate molecules in the microglia transcriptome that are induced by NT-3. This work will allow us to compile a database of potential molecules that may drive the CST sprouting response. In aim 2 we will determine whether NT-3 synergizes with microglia to enhance CST sprouting in vivo. Mouse genetics will be used to definitively prove a causal role for microglia in NT-3-induced axonal sprouting in vivo. Specifically, using Cre-lox technology, TrkC receptors will be selectively deleted in microglia. We predict that deleted TrkC in microglia will eliminate NT-3 re- programing of microglia and subsequently, their ability to promote axonal sprouting. In aim 3 we will use an acute and chronic rat unilateral pyramidotomy model to test, for the first time, if over-expression of NT-3 in the cervical spinal cord induces neuroplasticity and recovery of forelimb function. Additionally, we will test the effects of NT-3 over-expression in rats with cervical contusions. Th use of a more clinically-relevant vector and delivery method could lead to pre-translational research into the use of gene and cellular therapy for SCI. Practically, treatment for SCI must be effective after spontaneous functional recovery has plateaued, which could be months to years after the injury. Importantly, the studies proposed here target that chronic SCI window. Insights gained from these experiments have the potential to direct future therapeutic strategies to promote enhanced neuroplasticity and therefore increased functional recovery in patients with chronic SCI.